The type III secretion system of Bordetella bronchiseptica is chosen as a model system for studying bacterial-host interactions at the molecular level. B. bronchiseptica is a specialized respiratory pathogen closely related to B. pertussis (which causes whooping cough in humans). However, B. bronchiseptica naturally infects many laboratory animals, which allow us to study bacterial-host interactions in the context of natural infections. We have discovered a type III secretion system in B. bronchiseptica. Type III secretion systems are found in several pathogenic Gram-negative bacteria and can deliver proteins directly into the host cytoplasm or plasma membrane upon contact of the bacteria with the host cell. We have identified 22 linked genes in the B. bronchiseptica genome encoding proteins for a type III secretion system. By comparing the in vitro phenotypes of the wild type bacteria with a bscN deletion strain (that is defective in type III secretion) in their interactions with cultured cell lines, we deduced that type III secreted factors have a variety of effects on host cells. These include: induction of cytotoxicity, inhibition of activation of the transcription factor NF-kB by the aberrant aggregation of this factor in the cytoplasm, and activation of the ERK MAP kinase pathway. In vivo studies showed that type III secretion is required for persistent colonization in the trachea of rats and mice and the down-modulation of anti-Bordetella antibody production. Based on these observations, we hypothesize that type III secreted factors from B. bronchiseptica play immuno-modulatory roles for the bacteria to attain persistent, chronic colonization. [unreadable] [unreadable] The objective of this study is to identify the specific effector proteins secreted by the type III secretion system in B. bronchiseptica and determine how they alter host cellular processes and immune responses. We shall identify type III secreted proteins from B. bronchiseptica by both biochemical and genomic approaches, and use in vitro infection models to determine the mechanisms by which the effector protein(s) inhibit NF-kB activation and activate the MAP kinase pathways. We shall also use the mouse infection model to determine the importance of down-regulation of humoral immunity in allowing persistent colonization of the bacteria and how type III secreted virulence proteins interact with host immune cell functions. By identifying the bacterial effector proteins and examining how they interact with host cells, we shall determine the molecular basis for pathogenesis and down-regulation of immune processes.